Objective: PD-1/PD-L1 immune checkpoint treatment is effective for some triple-negative breast cancer populations with PD-L1 expression, but the response rate is still not satisfactory. This study aims to explore the mechanism of drug resistance to breast cancer anti-PD-1 therapies and the strategies for overcoming the resistance to PD-1therapies. Methods: By constructing a human triple-negative breast cancer drug-resistant cell line called BT-549R5 and a mouse breast cancer drug-resistant cell line called 4T1R3, and applying the whole-gene shRNA library screening, candidate drug resistance-associated molecules were obtained and verified by cytological experiments. The expression of Tyro3, Axl and MerTK of the TAM family in the 4T1R3 group was tested using the Western blot method. The down-regulation of CDK9 on the effect of T cells killing the BT-549R5 cells was observed through T cell killing tests, while the down-regulation of Tyro3 and CDK9 on the effect of anti-PD-1 therapies for transplanted breast tumors was observed in mouse tumor formation experiments. Results: The cell lines and animal models of breast cancer resistant to PD-1 treatment were successfully constructed. Tyro3, Axl and MerTK were highly expressed in 4T1R3 cells. Whole genome sequencing showed that Tyro3 and CDK9 were highly expressed in BT-549R5 cells. T cell killing experiment showed that the survival rate of BT-549R5 cells in the CDK9 down-regulated group and the control group decreased gradually with the increase of T cells, but the survival rate of BT-549R5 cells in the CDK9 down-regulated group decreased rapidly. Tumor formation experiment in mice showed that under anti-PD-1 treatment, the transplanted tumor in the 4T1R3 cell group grew rapidly compared with the 4T1 cell group (P<0.05), and the tumor volume of the 4T1R3 group was larger than that of the 4T1 group on Day 20. Nevertheless, the tumor growth rates in the CDK9-knockdown 4T1R3 cell group and the Tyro3-knockdown 4T1R3 cell group were similar to that of the 4T1 cell group, and the tumor volumes at day 20 were signiference lower than that of 4T1R3 cell group(P<0.05). Conclusions: Tyro3 and CDK9 are associated with the drug resistance to anti-PD-1 therapies for breast cancer. Inhibiting the expression of Tyro3 and CDK9 can reverse the drug resistance to breast cancer treatment.
Humans ; Animals ; Mice ; c-Mer Tyrosine Kinase/metabolism* ; Receptor Protein-Tyrosine Kinases/genetics* ; Axl Receptor Tyrosine Kinase ; Proto-Oncogene Proteins/metabolism* ; B7-H1 Antigen/genetics* ; Triple Negative Breast Neoplasms/genetics* ; Drug Resistance, Neoplasm ; Biomarkers ; Cell Line, Tumor ; Cyclin-Dependent Kinase 9

Objective: PD-1/PD-L1 immune checkpoint treatment is effective for some triple-negative breast cancer populations with PD-L1 expression, but the response rate is still not satisfactory. This study aims to explore the mechanism of drug resistance to breast cancer anti-PD-1 therapies and the strategies for overcoming the resistance to PD-1therapies. Methods: By constructing a human triple-negative breast cancer drug-resistant cell line called BT-549R5 and a mouse breast cancer drug-resistant cell line called 4T1R3, and applying the whole-gene shRNA library screening, candidate drug resistance-associated molecules were obtained and verified by cytological experiments. The expression of Tyro3, Axl and MerTK of the TAM family in the 4T1R3 group was tested using the Western blot method. The down-regulation of CDK9 on the effect of T cells killing the BT-549R5 cells was observed through T cell killing tests, while the down-regulation of Tyro3 and CDK9 on the effect of anti-PD-1 therapies for transplanted breast tumors was observed in mouse tumor formation experiments. Results: The cell lines and animal models of breast cancer resistant to PD-1 treatment were successfully constructed. Tyro3, Axl and MerTK were highly expressed in 4T1R3 cells. Whole genome sequencing showed that Tyro3 and CDK9 were highly expressed in BT-549R5 cells. T cell killing experiment showed that the survival rate of BT-549R5 cells in the CDK9 down-regulated group and the control group decreased gradually with the increase of T cells, but the survival rate of BT-549R5 cells in the CDK9 down-regulated group decreased rapidly. Tumor formation experiment in mice showed that under anti-PD-1 treatment, the transplanted tumor in the 4T1R3 cell group grew rapidly compared with the 4T1 cell group (P<0.05), and the tumor volume of the 4T1R3 group was larger than that of the 4T1 group on Day 20. Nevertheless, the tumor growth rates in the CDK9-knockdown 4T1R3 cell group and the Tyro3-knockdown 4T1R3 cell group were similar to that of the 4T1 cell group, and the tumor volumes at day 20 were signiference lower than that of 4T1R3 cell group(P<0.05). Conclusions: Tyro3 and CDK9 are associated with the drug resistance to anti-PD-1 therapies for breast cancer. Inhibiting the expression of Tyro3 and CDK9 can reverse the drug resistance to breast cancer treatment.
Humans ; Animals ; Mice ; c-Mer Tyrosine Kinase/metabolism* ; Receptor Protein-Tyrosine Kinases/genetics* ; Axl Receptor Tyrosine Kinase ; Proto-Oncogene Proteins/metabolism* ; B7-H1 Antigen/genetics* ; Triple Negative Breast Neoplasms/genetics* ; Drug Resistance, Neoplasm ; Biomarkers ; Cell Line, Tumor ; Cyclin-Dependent Kinase 9

BACKGROUND: As the morbidity and mortality in lung cancer keep raising, we are here to discuss the effect of clinical features especially the initial symptomon on diagnosis and follow-up treatment of newly diagnosed lung cancer patients. METHODS: The clinical features of the 500 patients with lung cancer in our hospital from March, 2017 to May, 2017 were analyzed retrospectively, including the initial symptom, stage, biomarkers, pathology, etc. RESULTS: There were 266 famle (53.3%), 372 adenocarcinoma (74.4%), 285 smokers (58%), status score of most patients (98.2%) was 0-1. 58.2% (n=291) of all the patients got biomarkers test, of which epidermal growth factor receptor (EGFR) mutations was 61.2%(178/291), anaplasticlymphoma kinase (ALK) fusion gene positive was 4.1% (12/291). Smoking status, initial symptom, pathological typing, TNM staging and EGFR mutation were the main factors affecting follow-up treatment. CONCLUSIONS Patients with typical symptoms have shorter diagnosis time. Smoking status, lung cancer-related symptoms, pathology, TNM staging and EGFR mutation status are the main factors that affect the follow-up treatment.
Adult ; Aged ; Aged, 80 and over ; Anaplastic Lymphoma Kinase ; China ; ErbB Receptors ; genetics ; metabolism ; Female ; Humans ; Lung Neoplasms ; diagnosis ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; Mutation ; Receptor Protein-Tyrosine Kinases ; genetics ; metabolism ; Retrospective Studies ; Smokers ; statistics & numerical data

Objective: To explore the effect of the AXL expression on the chemosensitivity of prostate cancer PC-3 and DU145 cells to docetaxel and possible mechanisms. METHODS: Using Western blot, we examined the expressions of the AXL protein, p-AXL and Gas6 in the docetaxel-resistant PC-3 (PC-3-DR) and DU145 (DU145-DR) cells stimulated with gradually increased concentrations of docetaxel. We transfected the PC-3 and DU145 cells with negative NC ShRNA and AXL-ShRNA, respectively, which were confirmed to be effective, detected the proliferation, apoptosis and cycle distribution of the cells by CCK8, MTT and flow cytometry after treated with the AXL-inhibitor MP470 and/or docetaxel, and determined the expression of the ABCB1 protein in the PC-3-DR and DU145-DR cells after intervention with the AXL-inhibitor R428 and/or docetaxel. RESULTS: The expression of the AXL protein in the PC-3 and DU145 cells was significantly increased after docetaxel treatment (P <0.05). The expressions AXL and p-AXL were remarkably higher (P <0.05) while that of Gas6 markedly lower (P <0.05) in the PC-3 and DU145 than in the PC-3-DR and DU145-DR cells. The inhibitory effect of docetaxel on the proliferation and its enhancing effect on the apoptosis of the PC-3 and DU145 cells were significantly decreased at 48 hours after AXL transfection (P <0.05). MP470 obviously suppressed the growth and promoted the apoptosis of the PC-3-DR and DU145-DR cells, with a higher percentage of the cells in the G2/M phase when combined with docetaxel than used alone (P <0.05). R428 markedly reduced the expression of ABCB1 in the PC-3-DR and DU145-DR cells, even more significantly in combination with docetaxel than used alone (P <0.05). CONCLUSIONS The elevated expression of AXL enhances the docetaxel-resistance of PC-3 and DU145 prostate cancer cells and AXL intervention improves their chemosensitivity to docetaxel, which may be associated with the increased cell apoptosis in the G2/M phase and decreased expression of ABCB1.
ATP Binding Cassette Transporter, Subfamily B, Member 1 ; metabolism ; Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Cell Count ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Docetaxel ; Drug Resistance, Neoplasm ; Humans ; Intercellular Signaling Peptides and Proteins ; metabolism ; Male ; Prostatic Neoplasms ; drug therapy ; metabolism ; pathology ; Proto-Oncogene Proteins ; drug effects ; genetics ; metabolism ; Pyrimidines ; pharmacology ; RNA, Small Interfering ; Receptor Protein-Tyrosine Kinases ; drug effects ; genetics ; metabolism ; Taxoids ; pharmacology

Axl encodes the tyrosine-protein kinase receptor, participating in the proliferation and migration of many cells. This study examined the role of Axl in functions of endothelial progenitor cells (EPCs). Axl was detected by RT-PCR and Western blotting in both placentas and EPCs from normal pregnancy and preeclampsia patients. The Axl inhibitor, BMS777-607, was used to inhibit the Axl signalling pathway in EPCs. Cell proliferation, differentiation, migration and adhesion were measured by CCK-8 assay, cell differentiation assay, Transwell assay, and cell adhesion assay, respectively. Results showed the expression levels of Axl mRNA and protein were significantly higher in both placentas and EPCs from preeclampsia patients than from normal pregnancy (P<0.05). After treatment with BMS777-607, proliferation, differentiation, migration and adhesion capability of EPCs were all significantly decreased. Our study suggests Axl may play a role in the function of EPCs, thereby involving in the pathogenesis of preeclampsia.
Adult ; Aminopyridines ; pharmacology ; Blood Pressure ; Case-Control Studies ; Cell Adhesion ; drug effects ; Cell Differentiation ; drug effects ; Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Female ; Fetal Blood ; cytology ; enzymology ; Gene Expression Regulation ; Gestational Age ; Human Umbilical Vein Endothelial Cells ; drug effects ; enzymology ; pathology ; Humans ; Placenta ; metabolism ; physiopathology ; Pre-Eclampsia ; blood ; genetics ; physiopathology ; Pregnancy ; Primary Cell Culture ; Protein Kinase Inhibitors ; pharmacology ; Proto-Oncogene Proteins ; antagonists & inhibitors ; genetics ; metabolism ; Pyridones ; pharmacology ; RNA, Messenger ; antagonists & inhibitors ; genetics ; metabolism ; Receptor Protein-Tyrosine Kinases ; antagonists & inhibitors ; genetics ; metabolism ; Stem Cells ; drug effects ; enzymology ; pathology

Accumulating studies explored the clinicopathologic and prognostic value of programmed death ligand-1 (PD-L1) in non-small cell lung cancer (NSCLC), but the results were controversial. We therefore conducted a meta-analysis to evaluate the predictive role of PD-L1 in NSCLC patients. We systematically collected relevant studies from PubMed, Embase, Web of Science and China National Knowledge Infrastructure. The pooled hazard ratios (HRs) with 95% confidence intervals (CIs) for overall survival (OS), and odd ratios (ORs) with 95% CIs for clinicopathologic factors were calculated. A total of 15 studies involving 3605 patients were included in this meta-analysis. The results showed no prognostic role of PD-L1 in the whole patients (HR=1.60, 95% CI: 0.88-2.89, P=0.123). Subgroup analysis showed that PD-L1 was associated with decreased OS in Asian patients (HR=2.00, 95% CI: 1.55-2.57, P<0.001). Among all the clinicopathologic factors, PD-L1 overexpression was significantly in relevance with poor tumor cell differentiation (HR=1.84, 95% CI: 1.49-2.28, P<0.001), late stage (HR=1.21, 95% CI: 1.02-1.43, P=0.026) and anaplastic lymphoma kinase (ALK) translocation (HR=2.63, 95% CI: 1.08-6.40, P=0.034), but not with other factors. In conclusion, our meta-analysis demonstrated that PD-L1 has a prognostic role in Asian patients with NSCLC.
Asian Continental Ancestry Group ; B7-H1 Antigen ; genetics ; metabolism ; Biomarkers, Tumor ; genetics ; metabolism ; Carcinoma, Non-Small-Cell Lung ; diagnosis ; ethnology ; genetics ; mortality ; European Continental Ancestry Group ; Humans ; Lung Neoplasms ; diagnosis ; ethnology ; genetics ; mortality ; Neoplasm Grading ; Neoplasm Staging ; Prognosis ; Proportional Hazards Models ; Protein Transport ; Receptor Protein-Tyrosine Kinases ; genetics ; metabolism

BACKGROUND: Statins are used to treat hypercholesterolemia; however, major cardiovascular events are decreased only 30% by statin treatment. Treatment with an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor has been reported to decrease serum glucose levels and improved insulin sensitivity in mice and humans, but there was no study in serum cholesterol levels. This study examined the effect of gefitinib, an EGFR tyrosine kinase inhibitor, on cholesterol metabolism in humans. METHODS: We retrospectively reviewed the medical records of 299 patients with primary lung cancer treated with gefitinib for ≥1 month and 72 patients with other treatments. Serum cholesterol, serum triglycerides, and body mass index were measured before and after treatment. The changes in serum cholesterol, serum triglycerides, and body mass index were compared between the gefitinib treatment group and the control group and were also analyzed according to the presence or absence of EGFR mutations. RESULTS: Serum cholesterol levels decreased significantly from 178.9 to 164.4 mg/dL after 1-month of gefitinib treatment. A total of 54 of the 299 patients underwent examination for the presence of the EGFR mutations. Serum cholesterol was significantly decreased in the group with the activating EGFR mutation (Δ=21.3 mg/dL) compared to that of those without the EGFR mutation (Δ=-3.1 mg/dL) after treatment with gefitinib. In contrast, there was no significantly difference between the two groups in control patients. CONCLUSION: Treatment with gefitinib decreased serum cholesterol in lung cancer patients, particularly in those with activating mutations in EGFR. These data suggest that EGFR tyrosine kinase inhibitors provide a novel and attractive strategy for the treatment of hypercholesterolemia.
Animals ; Blood Glucose ; Body Mass Index ; Cholesterol ; Epidermal Growth Factor ; Humans ; Hydroxymethylglutaryl-CoA Reductase Inhibitors ; Hypercholesterolemia ; Insulin Resistance ; Lung Neoplasms ; Lung ; Medical Records ; Metabolism ; Mice ; Protein-Tyrosine Kinases ; Receptor, Epidermal Growth Factor ; Retrospective Studies ; Triglycerides

Carcinoma, Non-Small-Cell Lung ; diagnosis ; metabolism ; therapy ; Humans ; Lung Neoplasms ; diagnosis ; metabolism ; therapy ; Receptor Protein-Tyrosine Kinases ; metabolism

CD151 is a member of the tetraspanin family that is implicated as a promoter of pathological or physiological angiogenesis. C-Met is expressed on a variety of cells including vascular endothelial cells (VECs) and up-regulated during angiogenesis. In this study, we investigated whether CD151 regulated migration, proliferation, tube formation and angiogenesis of human umbilical VECs (HUVECs) with activation of C-Met. Moreover, we studied whether CD151 could affect the angiogenic molecules such as nitric oxide (NO), vascular cell adhesion molecule-1 (VCAM-1) and vascular endothelial growth factor (VEGF). The expression of CD151 was determined by Western blotting. The cell proliferation assay was performed using the cell counting kit-8 (CCK-8) method and cell migration was assessed in microchemotaxis chambers by using fetal bovine serum (FBS) as the chemotactic stimulus. The angiogenic molecules were evaluated using ELISA. The NO level was detected using NO detection kit. The potential involvement of various signaling pathways was explored using relevant antibodies. We found that proliferation, migration and tube formation of HUVECs were promoted by CD151 with activation of C-Met, FAK and CDC42, while they were suppressed with CD151 knockdown by RNAi. Similarly, the levels of NO, VCAM-1 and VEGF in HUVECs were increased by CD151, but they were inhibited with CD151 knockdown by RNAi. These data suggested that CD151 could promote migration, proliferation, tube formation and angiogenesis of HUVECs, which was possibly related to the C-Met signaling pathways.
Base Sequence ; Human Umbilical Vein Endothelial Cells ; Humans ; Neovascularization, Physiologic ; RNA, Small Interfering ; genetics ; Receptor Protein-Tyrosine Kinases ; metabolism ; Signal Transduction ; Tetraspanin 24 ; genetics ; metabolism

OBJECTIVETo study the prevalence of ALK, ROS1 and RET fusion genes in non-small cell lung cancer (NSCLC), and its correlation with clinicopathologic features.

METHODSFormalin-fixed and paraffin-embedded tissue sections from samples of 302 patients with NSCLC were screened for ALK, ROS1, RET fusions by real-time polymerase chain reaction (PCR). All of the cases were validated by Sanger DNA sequencing. The relationship between ALK, ROS1, RET fusion genes and clinicopathologic features were analyzed.

RESULTSIn the cohort of 302 NSCLC samples, 3.97% (12/302) were found to contain ALK fusion genes, including 3 cases with E13; A20 gene fusion, 3 cases with E6; A20 gene fusion and 3 cases with E20; A20 gene fusion. There was no statistically significant difference in patient's gender, age, smoking history and histologic type. Moreover, in the 302 NSCLC samples studied, 3.97% (12/302) were found to contain ROS1 fusion genes, with CD74-ROS1 fusion identified in 9 cases. There was no statistically significant difference in patients' gender, age, smoking history and histologic type. One non-smoking elderly female patient with pulmonary adenocarcinoma had RET gene fusion. None of the cases studied had concurrent ALK, ROS1 and RET mutations.

CONCLUSIONSThe ALK, ROS1 and RET fusion gene mutation rates in NSCLC are low, they represent some specific molecular subtypes of NSCLC. Genetic testing has significant meaning to guide clinical targeted therapy.

Adenocarcinoma ; Aged ; Carcinoma, Non-Small-Cell Lung ; genetics ; metabolism ; Female ; Gene Fusion ; Genetic Testing ; Humans ; Lung Neoplasms ; Mutation ; Oncogene Proteins, Fusion ; genetics ; metabolism ; Protein-Tyrosine Kinases ; genetics ; metabolism ; Proto-Oncogene Proteins ; genetics ; metabolism ; Proto-Oncogene Proteins c-ret ; genetics ; metabolism ; Real-Time Polymerase Chain Reaction ; Receptor Protein-Tyrosine Kinases ; genetics ; metabolism ; Sequence Analysis, DNA ; Smoking